title: Splicing regulation of large exons secures phase-separation of transcription factors in vertebrates

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Ciliary associated extracellular vesicles modulate WNT signalling

status: Public on Jul 19 2021

city: Mainz

first: René

last: Ketting

address: Ackermannweg 4

country: Germany

institute: Institute of Molecular Biology

type: Non-coding RNA profiling by high throughput sequencing

summary: Primary cilia are microtubule based sensory organelles that protrude from almost every cell type. Their membrane contains highly specialized receptors important for receiving and processing extracellular signals, which enables them to regulate several signalling pathways. A recently discovered characteristic is that cilia are also able to release extracellular vesicles (EVs) from the ciliary membrane. Since EVs have been shown to exert numerous functions in physiology and pathology, these findings have the potential to dramatically alter our understanding of how the primary cilium is able to regulate various signalling pathways in development and disease. In our study we focused on the release of EVs from a ciliated kidney cell line. Using control and mutant cell lines, in which ciliary trafficking was disrupted, we observed that loss of primary cilia function leads to altered EV secretion and composition in NTA (Nanoparticle Tracking Analysis) and Western blot. Cilia mutant cells released more small EVs compared to control, and their composition was also changed between mutant and control. Protein identification via mass spectrometry identified both cilia- as well as WNT signalling-associated proteins and miRNA sequencing determined WNT-related miRNAs, which were differentially expressed in small EVs isolated from the cilia mutant cell line. Because of the presence of differentially expressed WNT related molecules in these small EVs, we tested whether this would have an effect on the WNT activity of recipient cells. We observed that small EVs secreted from cilia mutant cells differentially modulated the WNT response in recipient cells compared to control. Our results highlight a possible new small EV-dependent ciliary signalling mechanism, since deficient primary cilia lead to a change in EV secretion, resulting in an altered signal transduction. These results provide us with new insights into ciliopathy disease pathogenesis.

overall_design: EV preparations were obtained in triplicate from one control (WT) and two ciliary mutant (Bbs4 and Bbs6 knockout) cell lines. Cell lines were imortalised mouse kidney medullar cells as described in Hernandez-Hernandez V,  et al. (2013): Bardet-Biedl syndrome proteins control the cilia length through regulation of actin polymerization. Hum. Mol. Genet. 22(19):3858-68

first: Helen

middle: L

last: May-Simera

first: Ann-Kathrin

last: Volz

first: Eva-Maria

last: Albers

first: Karsten

last: Boldt

first: António

last: Miguel de Jesus Domingues

first: Rene


title: Grafting induced pluripotent stem cells-derived tenocytes contribute to recovery of motor function after Achilles tendon rupture in rat [single cell RNAseq]

city: Kyoto

first: Masayoshi

last: Tsukahara

state: Kyoto

address: Shogoin kawahara-cho, sakyo-ku,

department: Research and Develpoment center

country: Japan

institute: CiRA foundation

type: Expression profiling by high throughput sequencing

summary: Self-renewal of tendons is rare since the vascular formation inside is extremely poor, thus reconstructive surgery using autologous tendons has been often taken place in the case of severe injury. However, the rate of re-injury after surgery is relatively high, and collection of autologous tendons leads muscle weakness which results in prolonged rehabilitation. Here, we introduce the induced pluripotent stem cells (iPSCs)-based technology aiming at developing a new therapeutic option after tendon injury. We derived tenocytes from human iPSCs by recapitulating the normal progression of step-wise narrowing fate decisions in the vertebrate embryo. We used single-cell RNA sequencing to analyze the developmental trajectory of iPSCs-derived tenocytes.

overall_design: Single cell rnaseq analysis of the developmental trajectory of iPSC-derived tenocytes

first: Taiki

last: Nakajima

first: Tomoaki

middle: M

last: Kato

first: Makoto

last: Ikeya


title: Epigenetic programming of human sperm

status: Public on Jul 20 2021

city: Ste Anne-de-Bellevue

first: Romain

last: Lambrot

state: QC

address: 21111 Lakeshore road

department: Animal Science

country: Canada

institute: McGill University

type: Genome binding/occupancy profiling by high throughput sequencing

type: Methylation profiling by high throughput sequencing

summary:
The WGBS data was published in PMID: 31393794 and the sequencing data from WGBS and MCC-Seq have been submitted to the European Genome-phenome Archive under the accession number EGAS00001003617.
The paternal environment including stress, diet and toxicants has been linked to infertility and negative outcomes for offspring such as birth defects and adult onset of disease. Such effects may be transmitted via sperm through epigenetic mechanisms. To date, in depth profiling of the sperm epigenome in men has been limited. Our objective was to characterize the sperm profile of histone H3 lysine 4 tri-methylation (H3K4me3) from a reference population of men and relate this to sperm DNA methylation. ChIP-seq targeting H3K4me3 was performed on sperm from a representative reference population of 30 men and then overlapped with whole genome bisulfite sequencing (WGBS) data from the same men. Our analysis revealed that H3K4me3 is localized throughout the genome and at genes for fertility and development. Remarkably, enrichment was also found at regions that escape epigenetic reprogramming in primordial germ cells, embryonic enhancers and SINEs. The level of H3K4me3 in sperm associates with the degree of gene expression in embryo development. We find significant overlap in H3K4me3 and DNA methylation throughout the genome suggesting potential for the development of a personalized medicine approach for the assessment of fertility, lifestyle and environmental exposures.

overall_design:
Examination of H3K4me3 distribution in the sperm of a reference population of Canadian men.

Examination of H3K4me3 distribution (ChIP-seq) and DNA methylation (MCC-seq) in the sperm of Canadian men.

first: Sarah

last: Kimmins


title: Adipocyte-driven unfolded protein response is a shared transcriptomic signature of metastatic prostate carcinoma cells

status: Public on Jul 17 2021

city: Gainesville

first: Tracie

middle: R

last: Baker

state: FL

country: USA

institute: University of Florida

summary:
Purpose: To begin understanding molecular processes that enable tumor cells to survive and progress in difficult microenvironments such as bone, we performed unbiased examination of the transcriptome of two different prostate cancer cell lines in the absence or presence of bone marrow adipocytes.
Methods: 3’ RNA-seq (QuantSeq 3′ mRNA) was performed.  RNA from four biological replicates of PC3 and ARCaP(M) cells cultured alone or in Transwell with adipocytes was collected , and run on an Agilent TapeStation 2200 (Agilent Technologies, Santa Clara, CA) for quality control.  Lexogen’s QuantSeq 3’mRNA-seq Library Prep Kit (FWD for Illumina) was utilized for building RNAseq libraries.  The barcoded libraries were multiplexed at equimolar concentrations and sequenced with 50 bp single reads on an Illumina HiSeq-2500 run in rapid mode.  Data were demultiplexed using Illumina's CASAVA 1.8.2 software and reads were aligned to the human genomes. Differential gene expression analysis was used to detect adipocyte-mediated transcriptome changes.  qRT–PCR validation was performed using TaqMan  assays.
Results: Our RNAseq analyses and subsequent quantitative PCR and protein-based assays reveal that upregulation of Endoplasmic Reticulum (ER) stress and Unfolded Protein Response (UPR) genes is a shared signature between two metastatic prostate carcinoma cell lines of different origin. Pathway analyses and pharmacological examinations highlight the ER chaperone BIP as an upstream coordinator of this transcriptomic signature.
Conclusions: Our studies pave the way for additional mechanistic investigations and offer new clues towards effective therapeutic interventions in metastatic disease.

overall_design: RNA from four biological replicates of PC3 and ARCaP(M) cells cultured alone or in Transwell with adipocytes was sequenced

first: Izabela

last: Podgorski


title: Activation of a Hippocampal CREB-pCREB-miRNA-MEF2 Axis Modulates Individual Variation of Spatial Learning and Memory Capability

city: Taipei

first: I-Fang

last: Wang

address: Academia Rd.

department: Institute of Molecular Biology

country: Taiwan

institute: Academia Sinica

summary: To identify novel miRNAs involved in the regulation of learning and memory formation, we used the Morris water maze task to distinguish inbred wild-type C57BL/6J mice with good or poor learning and memory capability. C57BL/6J mice which undergo water maze task were sacrificed immediately after training. The hippocampus was harvested for miRNAs expression analysis with miRNA array (Phalanx Mouse & Rat miRNA OneArray® 2.0, annotation based on: miRBase release 15). These differentially expressed miRNAs were applied for further investigation.

overall_design: The hippocampus in experiment mice underwent Morris water maze with good performances or poor performances were harvested immediately after training (n=4 in each groups, total 2(Replicates)*2(groups)=4 arrays were used).

first: I

middle: F

first: Kuen

middle: J

last: Tsai

first: Che

middle: K

last: Shen


title: MiR-19a/miR-96-mediated low expression of KIF26A suppresses metastasis by regulating FAK pathway in gastric cancer

status: Public on Jul 16 2021

city: Jinan

first: Peng

last: Gao

address: Wen Hua Xi Road 44

country: China

institute: Shandong University

summary: Gastric cancer (GC) is one of the most common malignant neoplasms. Invasion and metastasis are the main causes of GC-related deaths. Recently, kinesins were discovered to be involved in tumor development. The aim of this study was to elucidate the roles of kinesin superfamily protein 26A (KIF26A) in GC and its underlying molecular mechanism in regulating tumor invasion and metastasis. Using real-time quantitative polymerase chain reaction (qPCR) and immunohistochemistry (IHC), we showed that KIF26A expression was lower in GC tissues without lymph node metastasis (LNM) than in nontumorous gastric mucosa, and even lower in GC tissues with LNM than in GC tissues without LNM. Functional experiments showed that KIF26A inhibited migration and invasion of GC cells. We further identifified focal-adhesion kinase (FAK), phosphatidylinositol 3-kinase regulatory subunit alpha (PI3KR1), VAV3, Rac1 and p21-activated kinase 2, and β-PAK (PAK3) as downstream effectors of KIF26A in the focal-adhesion pathway, and we found that KIF26A could regulate FAK mRNA expression through inhibiting c-MYC by MAPK pathway. c-MYC could bind to the promoter of FAK and activate FAK transcription. Moreover, we found that KIF26A mediated inactivation of the focal-adhesion pathway could reduce the occurrence of the epithelial-to-mesenchymal transition (EMT) by increasing expression of E-cadherin and reducing that of Snail. Luciferase assays and Western blotting revealed that miR-19a and miR-96 negatively regulate KIF26A. Finally, we found that decreased expression of KIF26A has been positively correlated with histological differentiation, Lauren classifification, LNM, distal metastasis, and clinical stage, as well as poor survival in patients with GC. These data indicate that KIF26A could inhibit GC migration and invasion by regulating the focal adhesion pathway and repressing the occurrence of EMT.

overall_design: We transfected GC SGC7901 cells with KIF26A siRNA 1 (siKIF26A 1) or NC for 48h. We then used the Whole Human Genome Oligo Microarray Assay (KangChen Bio-Tech, Shanghai, China) to analyze differential gene expression between the KIF26A siRNA and NCs groups.


title: mRNA-sequencing for HeLa cells expressing wild-type or mutant exogenous RPB6 and shRNA against endogenous RPB6

city: Yokohama

first: Yuki

last: Yamaguchi

state: Kanagawa

address: 4259 Nagatsuta-cho, Midori-ku

department: School of Life Science and Technology

institute: Tokyo Institute of Technology

summary: To elucidate the biological significance of the interaction between TFIIH-p62 and RPB6, we knocked down endogenous RPB6 expression and re-expressed wild-type (WT) or mutant (F13A, F8A-F13A, or ΔN20) RPB6 in HeLa cells using lentivirus vectors. While the doubling time of WT and F13A cells was close to that of parental cells (~24 h), the doubling time of F8A-F13A and ΔN20 cells was over 30 h. To investigate the underlying cause of this growth defect, we performed mRNA-sequencing of WT, F13A, F8A-F13A, and ΔN20 cells.

overall_design: We performed mRNA-sequencing for HeLa cells expressing wild-type (WT) or mutant (F13A, F8A-F13A, or ΔN20) exogenous RPB6 and shRNA against endogenous RPB6 in three biological replicates. Sequencing was performed using NextSeq 500 (Illumina).

first: Tetsufumi

last: Suwa


title: Lung dendritic cells migrate to the spleen to prime long-lived memory CD8+ T cell precursors after influenza virus infection

city: Birmingham

first: Alexander

last: Rosenberg

state: AL

address: 1720 2nd Avenue South, Suite 276/11

institute: University of Alabama at Birmingham

summary: CD8+ T cell responses to pulmonary challenges are primed by lung-migratory dendritic cells (mDCs), which capture antigens in the lung and migrate to the lung-draining mediastinal lymph node (med-LNs) to activate T cells. Notably, the lung and the spleen are not connected by the lymphatic vasculature. Thus, the current paradigm suggests that the med-LN is the only site for T cell priming to viruses that are restricted to the respiratory tract. Our results challenge this “LN-centric” paradigm. Using an influenza virus infection, we show here that lung-mDCs egress the med-LN and traffic to the spleen, where they prime influenza-specific CD8+ T cells. Importantly, CD8+ T cells primed in the spleen are transcriptionally different and have enhanced ability to differentiate into long-lived memory cells compared to med-LN-primed counterparts. Thus, our data reveal a previously ignored lung-mDC trafficking pathway that connects the lung with the spleen and has profound immunological implications.

overall_design: OTI cells (CD45.2) were adoptively transferred into day 1 PR8-infected CD45.1 mice. The next day, recipient mice were i.t treated with 60 μg of OVA and activated CD69+CD62Llo OTI cells were sorted from the spleen (spleen-primed, SP-P) and med-LN (med-LN-primed, LN-P) 24 later. Naïve OTI cells were also purified from naïve OTI mice and RNA-seq was performed.

first: Meagan

last: Jenkins

first: Holly

last: Bachus

first: Davide

last: Botta

first: Michael

middle: D

last: Schultz

first: Beatriz

last: León

first: André

last: Ballesteros-Tato


title: Serum mannose receptor promotes pro-inflammatory macrophage activation and obesity-associated metaflammation

city: Bonn

first: Joachim

last: Schultze

state: NRW

address: Carl-Troll-Strasse 31

department: Genomics and Immunoregulation

institute: LIMES (Life and Medical Sciences Center Genomics and Immunoregulation)

summary: Elevated serum concentrations of the soluble mannose receptor (sMR) have been reported to correlate with the severity of various inflammatory diseases. A physiological role of sMR, however, remains unclear. Here, we show that sMR binds CD45 on macrophages, both in vitro and in vivo, leading to cellular reprogramming towards an inflammatory phenotype by inhibition of CD45 phosphatase activity, which induces Src/Akt/NF-κB-mediated signaling. Remarkably, increased serum sMR levels correlate with obesity in both mice and humans. In addition, MR deficiency lowers the high-fat diet-induced increase in pro-inflammatory macrophages in metabolic tissues and protects against hepatic steatosis and whole-body metabolic dysfunctions. Conversely, administration of sMR induces serum pro-inflammatory cytokines and pro-inflammatory macrophages in the liver and promote insulin resistance. Altogether, our results reveal sMR as novel regulator of pro-inflammatory macrophage activation and could constitute a novel therapeutic target for hyperinflammatory diseases.

overall_design: To dissect the effect of the soluble Mannose Receptor (MR) on macrophages, we treated MR-deficient macrophages with a recombinant fusion protein between the Fc region of human IgG1 and the extracellular region of the MR (FcMR) for 4 h, 12 h or 24 h and isolated mRNA for RNA-seq analysis.

first: Jonas

last: Schulte-Schrepping

first: Maria

last: Embgenbroich

first: Kristian

last: Händler

first: Sven

last: Burgdorf


title: Genome-wide maps of RNA-protein interactions of SRSF3, hnRNPK and CPSF6

city: Nagoya

first: Akio

last: Masuda

state: Aichi

address: 65 tsurumai-cho, Showa-ku

institute: Nagoya University Graduate School of Medicine

summary: C2C12 cells were UV-irradiated at 400 mJ, and whole cell lysates were harvested from the cells. tRIP was performed as previously described (Masuda et al). Samples were sequenced on the Illumina NovaSeq6000 with 150 bp paired-end read (Macrogen, Japan) or Miseq with 150 bp single-read at the core facility of the Nagoya University. For paired-end read data, only P5 reads were used for analysis. Briefly, after standard HiSeq demultiplexing, reads were adapter-trimmed and reads less than 18 bp were discarded using cutadapt (v1.10). Mapping was first performed against the mouse repetitive elements in RepBase with STAR (v2.5.2b). Repeat-mapped reads were removed, and all others were then mapped against the mouse genome (mm10) with STAR (v 2.5.2b). Multiply mapped reads were filtered out. Duplicates of reads uniquely mapped to the human or mouse genome were removed by Picard (v2.0.1).

overall_design: Genome-wide maps of binding sites of RNA-binding proteins

first: Toshihiko

last: Kawachi


title: MicroRNAs in blood as biomarkers of colorectal cancer

city: Heidelberg

first: Andrea

middle: S

last: Bauer

address: Im Neuenheimer Feld 580

department: Functional Genome Analysis

institute: DKFZ

summary: Association studies have linked alterations of blood-derived circulating microRNAs (ci-miRNAs) with colorectal cancer (CRC). However, questions remain about the specificity of these abundance variations as biomarkers and their effects on the haematogenous spread of metastatic CRC (mCRC).

overall_design: We compared the profiles of miRNAs in 80 blood samples from healthy donors as well as patients with colorectal adenomas (CRA), colorectal diverticulitis (CRD), and CRC at different UICC-stages (I/II, III, and IV). We tested for stage-specific alterations

first: Axel

last: Stang


title: Expression analysis of Gluconacetobacter xylinus NBRC 3288 delta-fnrG mutant

city: Hanada-shi

first: Seiji

last: Watanabe

address: Nakamura-cho

department: Central Research Institute

institute: Mizkan Group Co. Ltd

summary: Investigation of whole genome gene expression level changes in a Gluconacetobacter xylinus NBRC 3288 delta-fnrG mutant, compared to the wild-type strain.

sample_organism: Komagataeibacter medellinensis NBRC 3288

overall_design: A 16 chip study using total RNA recovered from eight separate wild-type cultures of Gluconacetobacter xylinus NBRC 3288 and eight separate cultures of a disruptant mutant strain.  Each chip measures the expression level of 3,193 genes from Gluconacetobacter xylinus NBRC 3288 and the associated seven plasmid with twenty-one 60-mer probe pairs per gene, with three-fold technical redundancy.


title: RNA-sequencing of C. elegans long-lived mutants eat-2, ife-2, osm-5 and glp-1

city: Montreal

first: Jeremy

middle: Michael

last: Van Raamsdonk

address: 1001 Decarie Boulevard

department: Neurology and Neurosurgery

summary: Long-lived genetic mutants from different pathways of lifespan extension were used to determine the extent to which there are common downstream mediators of longevity. We have previously obtained RNA-sequencing data from other long-lived mutants including sod-2, clk-1, isp-1, nuo-6 and daf-2. Gene expression will be compared between these nine long-lived mutants.

overall_design: Six biological replicates per genotype. Worms were collected as prefertile young adults from NGM plates grown at 20 or 25 degrees Celsius with OP50 E. coli.

first: Meeta

last: Mistry


city: Oxford

first: Robin

last: Choudhury

address: L6 West Wing, John Radcliffe Hospital, Headley Way

department: Cardiovascular Medicine, Radcliffe Department of Medicine

country: United Kingdom

institute: University of Oxford

summary: Genome-wide histone modification profiling using ChIP-seq of diabetic or non-diabetic murine bone marrow derived macrophages (BMDM) and haematopoietic stem cells (HSC). Chromatin fragments were selected using anti-Histone H3 (tri methyl K4) (ab8580) or anti-histone H3 (acetyl K27) (ab4729). Sequencing libraries were prepared using the NEBNext Ultra II DNA Library Prep Kit.  instructions. The final libraries were purified using SPRI Ampure XP beads to remove adaptor dimers and sequenced by the Biomedical Sequencing Facility (BSF) at CeMM using the Illumina HiSeq 3000/4000 platform and the 50-bp single-read configuration.

overall_design: Wild-type C57BL/6J (12-14 weeks old) control or diabetic experimental groups. Diabetes was induced through an intra-peritoneal injection of streptozotocin (STZ) at a low dose range (42 – 45 mg / kg / day) over 5 consecutive days. After 6 weeks post-final STZ injection, diabetic mice with non-fasted blood glucose levels >13.9 mmol/L were sacrificed and tissue collected.

first: Laurienne

last: Edgar

first: Naveed

last: Akbar

first: Adam

middle: T

last: Braithwaite

middle: P


title: Oviduct epithelial cells constitute two developmentally distinct lineages that are spatially separated along the distal-proximal axis

status: Public on Jul 15 2021

city: montreal

first: Yojiro

last: Yamanaka

state: Quebec

address: 1160 Pine Avenue West, room 416

department: Human Genetics

institute: McGill University (Goodman Cancer Research Centre)

summary: Using a combination of scRNA sequencing, transgenic models and organoid culture we report the formation and maintenance of distinct distal and proximal epithelial populations in the mouse oviduct.

overall_design: Investigating heterogeneity in mouse oviduct epithelial cells

first: Matthew

last: Ford


title: Using transcriptomics and metabolomics to understand species differences in sensitivity to chlorpyrifos in Japanese quail and double-crested cormorant embryos

city: Ste anne de bellevue

first: Jessica

last: Head

address: 21111 Lakeshore Road

summary: Modern 21st century toxicity testing makes use of omics technologies to address critical questions in toxicology and chemical management. Of interest are questions relating to chemical mechanisms of toxicity, differences in species sensitivity, and translation of molecular effects to observable apical endpoints. Our study addressed these questions by comparing apical outcomes and multiple omics responses in early-life stage exposure studies with Japanese quail (JQ; Coturnix japonica) and double crested cormorant (DCCO; Phalacrocorax auritus), representing a model and ecological species, respectively. Specifically, we investigated the dose-dependent response of apical outcomes, transcriptomics, and metabolomics in the liver in each species exposed to chlorpyrifos (CPF), a widely used organophosphate pesticide. Our results revealed a clear pattern of dose-dependent disruption of gene expression and metabolic profiles in JQ but not DCCO at similar CPF exposure concentrations. The difference in effect sensitivity between species was likely due to higher metabolic transformation of CPF in the precocial JQ compared to the more altricial DCCO. The most impacted biological pathways after CPF exposure in JQ included hepatic metabolism, oxidative stress, endocrine disruption (steroid and non-steroid hormones), and metabolic disease (lipid and fatty acid metabolism). Importantly, we show consistent responses across biological scales, suggesting that significant disruption at the level of gene expression and metabolite profiles leads to observable apical responses at the organism level. Our study demonstrates the utility of evaluating effects at multiple biological levels of organization to understand how modern toxicity testing relates to outomes of regulatory relevance, while also highlighting important, yet poorly understood, species differences in sensitivity to chemical exposure.

sample_organism:
Phalacrocorax auritus
Coturnix japonica

platform_organism:
Phalacrocorax auritus
Coturnix japonica

overall_design: Controlled egg exposure study using two avian species, the japanese quail (JQ) and double crested cormorant (DDCO). Each species has a control (unexposed) and a medium and high dose exposure to chlorpyrifos. For JQ, the control, medium, and high doses have 5, 5, and 5 replicates, respectively. For DCCO, the control, medium, and high doses have 4, 5, and 5 replicates, respectively.

first: Jean-Pierre

middle: W

last: Desforges

first: Doug

last: Crump

first: Niladri

last: Basu


title: PRC1 drives Polycomb-mediated gene repression by controlling transcription initiation and burst frequency

first: Paula

last: Dobrinić

address: South Parks Road

department: Department of Biochemistry

summary: This SuperSeries is composed of the SubSeries listed below.

sample_organism:
Drosophila melanogaster
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: PAPERCLIP of cultured cells with/without DNA-damaging treatments

city: Pittsburgh

first: Hun-Way

last: Hwang

state: PA

address: 3550 Terrace Street

department: Department of Pathology

institute: University of Pittsburgh

summary: To identify APA changes in cancer cells in response to DNA damage, PAPERCLIP was performed on HeLa and LN229 cells with/without Doxorubicin treatment.

overall_design: PAPERCLIP was performed as previously described (Hwang et al. Cell Rep. 2016) and the libraries were sequenced on Illumina MiSeq and NextSeq.


title: Effect of lactoferrin treatment on iAEC2

city: Ann Arbor

first: Arul

middle: M.

last: Chinnaiyan

state: MICHIGAN

address: 1500 E. Medical Center Dr

institute: University of Michigan

summary: In an attempt to understand the mode of antiviral action of lactoferrin on SARS-CoV-2 infection, we focused on the effect of lactoferrin on uninfected cells. We decided to use a physiologically relevant cell model of iPSC derived- alveolar epithelial cells (the iAEC2). We specifically focused on antiviral pathway triggered by lactoferrin on treated cells that could explain a cell-mediated effect of lactoferrin on viral replication

overall_design: We seeded iAEC2 cells in collagen-treated plate and allowed them to adhere. When the cells reached confluence, they were treated with lactoferrin at a concentration of 6.25uM (500ug/ml) for two days. Cells were then harvested in TriZol, RNA was extracted with the DirectZol kit (Zymogen) according to manufacturers instructions and was sequences with the Illumina NovaSeq 6000 System.

first: Carmen

last: Mirabelli

first: Jesse

last: Wotring

first: Charles

last: Zhang

first: Sean

last: McCarty

first: Reid

last: Fursmidt

first: Carla

last: Pretto

first: Yuanyuan

last: Qiao

first: Yuping

first: Tristan

last: Frum

first: Namrata

last: Kadambi

first: Anya

last: Amin

first: Teresa

last: O’Meara

first: Jason

last: Spence

first: Jessie

last: Huang

first: Konstantinos

last: Alysandratos

first: Darrell

middle: N

last: Kotton

first: Samuel

last: Handelman

first: Christiane

middle: E

last: Wobus

first: Kevin

last: Weatherwax

first: George

middle: A

last: Mashour

first: Jonathan

middle: Z

last: Sexton


title: rs41291957 alters secondary RNA structure of the human pri-miR-143/145

city: Brescia

first: Leonardo

last: Elia

state: Brescia

address: v.le Europa 11

department: Dipartimento di Medicina Molecolare e Traslazionale

country: Italy

institute: Università degli Studi di Brescia

summary:
Purpose: To evaluate whether the single nucleotide polimorphsim rs41291957, located in the pri-miR-143/145, influences the primary RNA secondary structure.
Methods: In vitro transcription and folding  for the pri-miR-143/145 carrying the WT allele (G) or the mutated one (A) was carried out. The RNAs were then subjected to RNAseI treatement for 30 minutes and RNA seq performed on the digested nucleic acids.
Results: Using an optimized data analysis workflow for small RNAs, we mapped the sequence reads on the human pri-miR-143/145 carrying the G- or A-allele. We observed an increase of small reads (<60bps) for the A-allele and a different profile of read enrichement between the digested G- and A-allele RNA, indicating the difference in secondary structure.

overall_design: Examination of 2 different pri-miR-143/145, carrying eaither the G- or A-allele.

first: Montserrat

last: Climent

first: Luca

last: Lambroia


title: Transcriptomic analysis of intestinal biopsies in Zambian adults with environmental enteropathy

city: London

first: Kelly

last: Pauk

address: Turner Street

department: Blizard Institute

institute: Queen Mary University of London

summary: Study of adults with environmental enteropathy from residential area of Lusaka, Zambia

overall_design: Cross-sectional study of RNAseq data.

first: Paul

last: Kelly

first: Mubanga

last: Chama


title: Drosophila architectural protein CTCF is not essential for fly survival and is able to function independently of CP190

city: Moscow

first: Natalia

last: Klimenko

address: 34/5 Vavilova Street

country: Russia

institute: Institute of Gene Biology (IGB) of the Russian Academy of Sciences

summary: CTCF is the most likely ancestor of proteins that contain large clusters of C2H2 zinc finger domains (C2H2) and is conserved among most bilateral organisms. In mammals, CTCF functions as the main architectural protein involved in the organization of topology-associated domains (TADs). In vertebrates and Drosophila, CTCF is involved in the regulation of homeotic genes. Previously, null mutations in the dCTCF gene were found to result in death during the stage of pharate adults, which failed to eclose from their pupal case. Here, we obtained several new null dCTCF mutations and found that the complete inactivation of dCTCF appears to be limited to phenotypic manifestations of the Abd-B gene and fertility of adult flies. Many modifiers that are not associated with an independent phenotypic manifestation can significantly enhance the expressivity of the null dCTCF mutations, indicating that other architectural proteins are able to functionally compensate for dCTCF inactivation in Drosophila. We also mapped the 715–735 aa region of dCTCF as being essential for the interaction with the BTB (Broad-Complex, Tramtrack, and Bric a brac) and microtubule-targeting (M) domains of the CP190 protein, which binds to many architectural proteins. However, the mutational analysis showed that the interaction with CP190 was not essential for the functional activity of dCTCF in vivo.

overall_design: ChIP-seq signal of the HA, dCTCF and CP190 architectural/insulator proteins occupancy in whole adult flies in dCTCF wt and dCTCFΔ705-733 lines.

first: O

last: Maksimenko

first: N


title: RNA-seq of diabetic and case control human peripheral blood mononuclear cells (PBMC)

first: Robin

last: Choudhury

summary: Purpose: RNA-sequencing of diabetic and case control human peripheral blood mononuclear cells (PBMCS) to determine differentially expressed genes and pathways in control-unstimulated and M1 proinflammatory stimulated cells for determination of hyperglycaemic memory.

first: Laurienne

last: Edgar

first: Naveed

last: Akbar

first: Adam

last: Braithwaite


title: Next Generation Sequencing Facilitates Quantitative Analysis of Adipose Tregs from Foxp3Cre and Hif1afl/fl;Foxp3Cre mice

city: Shanghai

first: Bin

last: Li

address: South Chongqing Road 208

institute: Shanghai Institue of Immunology

summary: The goal of this study is to study mechanisms underlying how HIF-1α regulates adipose Treg function. Therefore, mRNA profiles of adipose Tregs (from 4-month old Foxp3Cre and Hif1afl/fl;Foxp3Cre mice) were generated by deep sequencing, single experiment, using Illumina HiSeq TEN. We compared gene expression profiles of adipose Tregs from Hif1afl/fl;Foxp3Cre and Foxp3Cre mice. Using unbiased comparative gene expression analyses, we found adipose HIF-1α-ΔTreg cells showed higher expression of Nt5e gene and simultaneously down-regulated canonical adipose Treg genes (including Pparg, Il1rl1, Klrg1, Areg). Therefore, these data suggested a dependency of HIF-1α on PPAR-γ expression in adipose Tregs.

overall_design: mRNA profiles of adipose Tregs (from 4-month-old Foxp3Cre and Hif1afl/fl;Foxp3Cre mice) were generated by deep sequencing, single experiment, using Illumina HiSeq TEN.

first: Yangyang


title: Gene Expression Analysis of Blood PBMC Response to Mycobacterium leprae antigens

city: Zhengzhou

first: Youhua

last: Yuan

state: Henan

address: The People's Hospital, Zhengzhou University

department: Clinical Microbiology

institute: Henan Provincial People's Hospital

summary:
8 leprosy patients including 4 multibacillary (MB) and 4 paucibacillary (PB), and 8 non-leprosy controls including 4 healthy house contacts (HHCs) and
4 endemic controls (ECs) were included in the study. The immune response differences between leprosy patients and controls were evaluated by analyzing the transcriptional profiles of PBMCs to M. leprae sonicate antigens by RNA-seq. The analyses revealed potential biomarkers (including mRNAs and lncRNAs) preferentially expressed in PBMCs in leprosy patients that may be useful for early diagnosis of leprosy.

overall_design: PBMCs were isolated from leprosy patients and controls, then stimulated with M. leprae whole cell sonicate antigens (10 µg/ml) followed by RNA-seq analysis.

first: Y

first: J

last: Liu

last: You

first: X

last: Chen

first: L

last: Wen

first: H

last: Zhang


title: Diet-induced alteration of intestinal stem cell identity and lineage allocation underlies obesity and pre-diabetes in mice

city: Neuherberg

first: Heiko

last: Lickert

address: Ingolstaedter Landstraße 1

department: Institute of Diabetes and Regeneration Research

institute: Helmholtz Zentrum München German Research Center for Environmental Health

summary: Excess nutrient uptake and altered hormone secretion in the gut contribute to a systemic energy imbalance causing obesity and increased risk for type 2 diabetes (T2D) and colorectal cancer. This functional maladaptation is thought to emerge at the level of the intestinal stem cells (ISCs). But, how an obesogenic diet affects ISC identity and fate decisions is not well understood. Here, we study the intestinal responses to an obesogenic diet in mice by combining single-cell profiling of more than 27,000 small intestinal crypt cells with genetic lineage labelling and tracing of ISC fate decisions and in situ metabolomics. We find that an obesogenic diet induces ISC and progenitor hyperproliferation, but also enhances ISC differentiation and cell turnover. Furthermore, the regional identity of ISCs and enterocytes is reprogrammed from a distal to a proximal phenotype and enterocyte zonation is altered to adapt cell function to nutrient availability. Single-cell resolution of the enteroendocrine lineage shows an increase in endocrine progenitors and peptidergic enteroendocrine cell (EEC) types and a decrease in serotonergic EEC types. Mechanistically, we can link increased fatty acid synthesis, Ppar signalling and the Insr/Igf1r/Akt pathway to ISC dysfunction, hyperproliferation and impaired endocrine differentiation. In summary, our work describes key molecular mechanisms of diet-induced intestinal maladaptation in mice that promote obesity, and, thus, underlie the pathogenesis of the metabolic syndrome and associated complications.

overall_design: We performed Affymetrix microarray analysis on FACS purified intestinal crypt cells expressing different levels of Foxa2-Venus Fusion (FVF) protein (FVF-negative, FVF-low, FVF-high) isolated mice fed a high-fat-high-sugar or control diet.

first: Alexandra

last: Aliluev

first: Sophie

last: Tritschler

first: Michael

last: Sterr

first: Lena

last: Oppenländer

first: Julia

last: Hinterdobler

first: Tobias

last: Greisle

first: Martin

last: Irmler

first: Johannes

last: Beckers

last: Sun

first: Axel

last: Walch

first: Kerstin

last: Stemmer

first: Alida

last: Kindt

first: Jan

last: Krumsiek

first: Matthias

middle: H

last: Tschöp

first: Malte

last: Lücken

first: Fabian

last: Theis

first: Anika

last: Böttcher


title: Single-cell ATAC-Sequencing of adipose Treg cells

first: Bin

last: Li

summary: Systematic characterizations of adipose Treg subsets and their in vivo roles remain uncommon. The goal of this study is to reveal the heterogeneity and chromatin landscape of adipose Treg cells. Using single-cell ATAC-seq to map adipose Treg cells, we identified different adipose subsets. Strikingly, our scATAC-seq analysis revealed a strong enrichment of key transcription factor motifs, which may play function in fate specification. By gain and loss of function studies, we further demonstrated adipose Treg subsets display disparate physiologic influences. Our findings have important implications for understanding phenotypic diversity of tissue Tregs in vivo.

overall_design: Around 60,000 TCRβ+CD4+CD25+YFP+ adipose Treg cells from 4-month old male Foxp3Cre mice were subjected to nuclei isolation according to the demonstrated protocol: Nuclei Isolation for Single Cell ATAC Sequencing (10x Genomics). scATAC-seq libraries were prepared according to the Chromium Single Cell ATAC Reagent Kits User Guide (10x Genomics; CG000168 Rev A).

first: Yangyang


title: Defining the nature of human pluripotent stem cell-derived interneurons via single cell analysis

city: Los Angeles

first: Kathrin

last: Plath

state: CA

address: 615 Charles E. Young Drive South

institute: University of California, Los Angeles

summary: We applied single nuclei and single cell transcriptomics methods to model interneuron development with interneurons derived from human PSCs. We find that PSC-derived interneurons capture stages of development prior to mid gestation, but only represent a minority of potential subtypes that are found in the brain. Comparison of these interneurons to those found in the adult brain highlighted decreased expression of synapse-related genes, reflective of their electrophysiological immaturity. Together, these analyses provide a full characterization of PSC-derived interneurons.

overall_design: hESCs cultures underwent directed differentiation into interneurons, and were then isolated for single cell RNA-seq.

first: Thomas

last: Allison

first: Justin

last: Langerman

first: Shan

last: Sabri

first: Marcos

last: Otero-Garcia

first: Andrew

last: Lund

first: John

last: Huang

first: Xiaofei

last: Wei

first: Ranmal

last: Samarsinghe

first: Damon

last: Pouladakis

first: Istvan

last: Mody

first: Inma

last: Cobos

first: Bennett

last: Novitch

first: Daniel

last: Geschwind

first: William

last: Lowry


title: Plant growth and salt stress responses

city: West Lafayette

first: Jian-Kang

last: Zhu

state: IN

address: 625 Agriculture Mall Dr.

department: Department of Horticulture and Landscape Architecture

institute: Purdue University


title: The LRX-RALF-FER cell wall integrity sensing module controls plant growth and salt stress responses by modulating multiple plant hormones

summary: Plants have evolved cell wall integrity signaling pathways to maintain cell wall homeostasis during rapid growth and in response to environmental stress. The cell wall leucine-rich repeat extensins LRX3/4/5, the RAPID ALKALINIZATION FACTOR (RALF) peptides RALF22/23, and FERONIA (FER) function as a module to regulate plant growth and salt stress responses via the sense of cell wall integrity. However, the intracellular signaling pathways that mediate the effects of the LRX3/4/5-RALF22/23-FER module are still largely unknown. Here, we report that jasmonic acid (JA), salicylic acid (SA), and abscisic acid (ABA) accumulate constitutively in lrx345 and fer mutants. Blocking JA pathway rescues the retarded growth phenotype of the lrx345 and fer-4 mutants, while disruption of ABA biosynthesis suppresses the salt-hypersensitivity of these mutants. Many salt stress-responsive genes display abnormal expression patterns in the lrx345 and fer-4 mutants, as well as in wild type plants treated with epigallocatechin gallate (EGCG), an inhibitor of pectin methylesterases, suggesting that the cell wall integrity is a critical factor that determines the expression of stress-responsive genes. Production of reactive oxygen species (ROS) is constitutively increased in the lrx345 and fer-4 mutants, and inhibition of ROS accumulation suppresses the salt-hypersensitivity of these mutants. Together, our results suggest that the LRX3/4/5-RALF22/23-FER module controls plant growth and salt stress responses by regulating hormonal homeostasis and ROS accumulation.

overall_design: RNA-Seq of 8 samples. The samples are wild type ( WT ) and lrx345 after NaCl treatment for 0, 6, 24, and 72 h.

first: Chunzhao

last: Zhao

first: Kai

last: Tang


title: Cut-and-run sequencing of adipose Treg cells with anti-HIF-1α and anti-PPAR-γ antibodies

first: Bin

last: Li

summary: Molecular mechanisms underlying the development of adipose Tregs remain poorly understood, prompting us to subsequently characterize the critical transcription cascade. The goal of this study is to reveal the function of key transcription factors HIF-1α and PPAR-γ in fate specification of adipose Tregs. Using CUT&RUN assay and sequencing of the enriched DNA, we detected a binding of HIF-1α and PPAR-γ to specific genomic regions in adipose Treg cells. In doing so, we highlighted a potential role of HIF-1α and PPAR-γ in establishing the specific identity of adipose Tregs.

overall_design: Around 30,000 TCRβ+CD4+CD25+YFP+ adipose Treg cells from 4-month old male Foxp3Cre mice were subjected to CUT&RUN assay according to the demonstrated protocol (CUT&RUN, Cell Signaling Technology, Cat#14209). Extracted DNA was subjected to sequencing.

first: Yangyang


title: A regulatory module controls plant growth and stress response by modulating hormone level

summary: Perception and relay of cell wall signals is critical for plants to regulate growth and stress responses, but the mechanism underlying this biological process remains largely unknown. Here we report that RAPID ALKALINIZATION FACTOR (RALFs) peptides, Leucine-rich repeat extensins (LRXs) and FERONIA (FER) are physically associated in extracellular region. lrx345, fer-4, and transgenic plants overexpressing RALF22 or RALF23 all showed retarded growth, increased sensitivity to salt stress, and constitutively increased levels of ABA, JA, and SA. Salt treatment or chemical disruption of cell wall promotes the release of mature RALF peptides, which negatively regulate the function of FER by inducing its internalization. JA mutants coi1 and aos restore the retarded growth, and mutation of ABA2 suppresses the salt-sensitive phenotype of lrx345. Together, we propose that LRXs, RALFs, and FER function as a module to sense and transduce cell wall signals, thereby regulating growth and stress responses via hormones-mediated pathways.

overall_design: RNA-Seq of 4 samples, each has 3 replicates. The samples are wild type ( WT ) and lrx345 before and after NaCl treatment.


title: RNA-seq of glioma and normal cell lines after HDAC1 knock-down

status: Public on Jul 14 2021

city: Tempe

first: Christopher

middle: L

last: Plaisier

state: Arizona

department: SBHSE

institute: Arizona State University

summary: Using RNA-seq we characterized the effect of shRNA mediated knockdown of HDAC1 in glioblastoma stem cells (GSCs), normal ihNPCs, and normal human astrocytes.

overall_design: GSCs, hNPCs and NHAs transduced with either shNT or shHDAC1 shRNAs at a MOI of 3 were selected with 2 ug/mL puromycin 72 hours post-transduction. 72 hours post-selection, total RNA was extracted from cells using the PureLink RNA Mini Kit (Ambion) in accordance to the manufacturer's instructions.

first: Costanza

last: Lo Casio

first: James

last: McNamara

first: Ernesto

last: Melendez

first: Erika

last: Lewis

first: Matthew

last: Dufault

first: Harley

middle: I

last: Kornblum

first: Nader

last: Sanai

first: Shwetal

last: Mehta


title: Inferring regulatory targets of transcription factors from simultaneous single cell measurements of intranuclear proteins and gene expression

city: Cambridge

first: Hattie

last: Chung

state: MA

address: 415 Main St

institute: Broad Institute

summary: Simultaneous single cell measurements of nuclear proteins and RNA with inCITE-seq, with targeted protein measurements using DNA-conjugated antibodies. Single nuclei (n=10,014) of HeLa cells, either untreated or after TNFa treatment, were profiled with inCITE-seq targeting the transcription factor p65. Single nuclei (n=21,583) of the mouse hippocampus after PBS or kainic acid injection were profiled with inCITE-seq targeting TFs p65, c-Fos, and PU.1, as well as the RNA-binding protein NeuN.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Joint single cell measurements of targeted nuclear proteins and RNA with inCITE-seq.

first: Aviv

last: Regev


title: Targeting Histone K4 Trimethylation for Treatment of Cognitive and Synaptic Deficits of Mouse Models of Alzheimer's Disease (RNA-seq)

city: Buffalo

first: Zhen

last: Yan

state: NY

address: 955 Main St

institute: SUNY Buffalo

summary: We performed RNAseq experiments to examine genome-wide transcriptional changes in PFC of P301S Tau mice with or without WDR5-0103 treatment.

overall_design: mRNA via RNAseq analysis of two 6 mo. old  PS19(P301S), and two 6 mo. old control (C57BL/6 x C3H), and two 6 mo. old PS19P301S) treated with WDR5-0103  mice from PFC tissue were compared.

first: Jamal

last: Williams

first: Qing

last: Cao


title: UTX condensation underlies its tumor suppressive activity

city: Charlottesville

first: Bi

last: Shi

state: Virginia

address: 1340 JPA Pinn Hall Room 6017

department: Department of Biochemistry and Molecular Genetics

institute: University of Virginia

summary: We investigated whether UTX condensates regulate local H3K4me1 in the genome by ChIP-seq of THP-1 cells expressing empty vector (control), UTX WT, ΔcIDR, UTX-eIFIDR, or ΔTPR. To determine the effect of UTX condensation on high-order chromatin interactions or looping, we performed H3K4me3 HiChIP in duplicates to generate high-resolution contact maps around the active and poised transcription start sites (TSSs) in THP-1 cells expressing empty vector (control), UTX WT, ΔcIDR, or UTX-eIFIDR.

overall_design: ChIP was performed and libraries were constructed and sequenced with 20 million THP-1 cells expressing empty vector (control), UTX WT, ΔcIDR, UTX-eIFIDR, or ΔTPR.   HiChIP experiments were performed with 2.5 million THP-1 cells expressing empty vector (control), UTX WT, ΔcIDR, UTX-eIFIDR, or ΔTPR using the Arima-HiC+ kit according to its recommended protocol.

first: Zhenjia

last: Wang

first: Chongzhi

last: Zang

first: Hao

last: Jiang


title: Genome-wide mapping of SETMAR transposase binding sites in the human genome

city: Debrecen

first: Márton

last: Miskei

address: Egyetem tér 1.

department: Department of Molecular Biotechnology and Microbiology

country: Hungary

institute: University of Debrecen

summary: SETMAR/Metnase is a naturally occurring fusion protein that consists of a histone-lysine methyltransferase domain and an HsMar1 transposase. To elucidate the biological role of SETMAR, it is crucial to identify genomic targets to which SETMAR specifically binds and link these sites to the regulation of gene expression. Herein, we mapped the genomic landscape of SETMAR in a near-haploid human leukemia cell line (HAP1) in order to identify on-target and off-target binding sites at high resolution and to elucidate their role in terms of gene expression. Our analysis revealed a perfect correlation between SETMAR and inverted tandem repeats (ITRs) of HsMar1 transposon remnants, which are considered as natural target sites for SETMAR chromosome binding. However, we did not detect any untargeted events at non-ITR sequences, calling into question previously proposed off-target binding sites. We identified sequence fidelity of the ITR motif as a key factor for determining the binding affinity of SETMAR for chromosomes, as higher ITR fidelity resulted in increased affinity for chromatin and stronger repression of SETMAR-bound gene loci. These associations highlight how SETMAR’s chromatin binding fine-tune gene regulatory networks in human tumour cells.

overall_design: ChIP-seq analysis on HAP1 haploid human cell line to map genomic landscape of SETMAR.

first: Marton

first: Lorant

last: Szekvolgyi


title: U2af1 is required for survival and function of hematopoietic stem/progenitor cells

first: Golam

last: Mohi

address: 1340 Jefferson Park Ave

department: Biochemistry and Molecular Genetics

summary: U2AF1 is involved in the recognition of the 3’ splice site during pre-mRNA splicing. Mutations in U2AF1 are frequently observed in myelodysplastic syndromes. However, the role of wild-type U2AF1 in normal hematopoiesis has remained elusive. Using a novel conditional U2af1 knockout allele, we have found that deletion of U2af1 results in profound defects in hematopoiesis characterized by pancytopenia, ablation of hematopoietic stem/progenitor cells (HSPC) leading to bone marrow failure and early lethality in mice. U2af1 deletion impairs HSPC function and repopulation capacity. U2af1 deletion also causes increased DNA damage and reduced survival in hematopoietic progenitors. RNA sequencing analysis reveals significant alterations in the expression of genes related to HSC maintenance, cell proliferation and DNA damage response-related pathways in U2af1-deficient HSPC. U2af1 deficiency also induces splicing alterations in genes important for HSPC function. Collectively, these results suggest an important role for U2af1 in the maintenance and function of HSPC in normal hematopoiesis. A better understanding of the normal function of U2AF1 in hematopoiesis is important for development of appropriate therapeutic approaches for U2AF1 mutant induced hematologic malignancies.

overall_design: Transcriptome and splicing alterations in U2af1-deficient mouse bone marrow LSK and lineage-negative cells

first: Avik

last: Dutta

first: Bao

last: Le


title: Single-cell Sequencing of adipose Treg cells

first: Bin

last: Li

summary: Systematic characterizations of adipose Treg subsets and their in vivo roles remain uncommon. Using paired single-cell RNA-seq and TCR-seq to map adipose Treg cells, we identified conserved adipose subsets with distinct clonal expansion patterns. By gain and loss of function studies, we demonstrated adipose Treg subsets display disparate influences. To understand the clonal structure of adipose Treg cells, we analyzed their TCR sharing data and observed a number of overlapped TCR clonotypes, suggesting a cellular state transition between adipose Treg clusters. Our findings have important implications for understanding phenotypic diversity of tissue Tregs in vivo.

overall_design: Around 30,000 TCRβ+CD4+CD25+YFP+ adipose Treg cells from Foxp3Cre mice were sorted by flow cytometry into a chilled tube filled with 30μl PBS with 0.04% BSA. Cells were then encapsulated in one lane of a 10x Chromium instrument, Libraries were constructed with a Single Cell 5′ Reagent Kit. And TCR transcripts can be enriched with a Single Cell V(D)J Reagent Kit. We obtained 14,274 good quality cells after filtering. We first generated a resource on the gene expression profiles of murine adipose Treg cells.

first: Yangyang


title: NUT Carcinoma

city: Boston

first: Hitoshi

last: Shiota

state: Massachusetts

address: 77 Avenue Louis Pasteur, NRB rm652

institute: Brigham and Women's Hospital

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Expression of Foxp3 by T follicular helper cells in end-stage germinal centers

city: New York

first: Johanne

last: Jacobsen

address: 1230 York Avenue

institute: The Rockefeller University

summary: Germinal centers (GCs) are the site of immunoglobulin somatic hypermutation and affinity maturation, processes essential to an effective antibody response. The formation of GCs has been studied in detail, but less is known about what leads to their regression and eventual termination, factors that ultimately limit the extent to which antibodies mature within a single reaction. We show that contraction of immunization-induced GCs is immediately preceded by an acute surge in GC-resident Foxp3+ T cells, attributed at least partly to upregulation of the transcription factor Foxp3 by T follicular helper (Tfh) cells. Ectopic expression of Foxp3 in Tfh cells is sufficient to decrease GC size, implicating the natural upregulation of Foxp3 by Tfh cells as a potential regulator of GC lifetimes.

overall_design: Single cell RNA-Seq of germinal center T cells and single cell RNA-Seq of germinal center T cells upon ectopic Foxp3 expression.

first: Wei

last: Hu

first: Tiago

middle: B

last: Castro

first: Sigrid

last: Solem

first: Alice

last: Galante

first: Zeran

last: Lin

first: Samuel

last: Allon

first: Luka

last: Mesin

first: Angelina

middle: M

last: Bilate

first: Ariën

last: Schiepers

first: Alex

last: Shalek

first: Alexander

middle: Y

last: Rudensky

first: Gabriel

last: Victora


title: ChIP-seq analysis of BRD4-NUT and H3K27ac distribution in NUT Carcinoma cell line TC-797 treated with DMSO or Panobinostat

summary: BRD4-NUT and H3K27ac are depleted from megadomains in NUT Carcinoma (NC) cell line TC-797 after treatment with Panobinostat.

overall_design: ChIP-seq analysis of BRD4-NUT and H3K27ac distribution in NC cell line TC-797 treated with DMSO or Panobinostat.

first: Madeleine

last: Lemieux

first: Christopher

last: French


title: Next Generation Sequencing Facilitates Quantitative Analysis of adipose tissue Treg and LN Treg Transcriptomes

first: Bin

last: Li

summary: The goal of this study is to reveal unique features of adipose Tregs and to study mechanisms underlying how mediator Med23 regulate adipose Treg function in aged mice. Therefore, mRNA profiles of LN Tregs (from 4-month old Foxp3Cre mice) and adipose Tregs (from 4-month old Foxp3Cre, 4-month old obese Foxp3Cre, 10-month old Foxp3Cre and 10-month old Med23fl/fl;Foxp3Cre mice) were generated by deep sequencing, single experiment, using Illumina HiSeq2000. We compared transcriptome features between lymph node-derived Tregs and adipose Tregs from 4-month old lean or obese mice. Using unbiased comparative gene expression analyses, we found adipose Tregs display an up-signature of Insr (Insulin receptor) and Hif1a, while Pparg acts as a positive control. We next compared gene expression profiles of adipose Tregs from 10-month old Med23fl/fl;Foxp3Cre (MKO) and Foxp3Cre (WT) mice. adipose Med23-ΔTreg cells display impaired transcription of Pparg and Il1rl1 (ST2), and they simultaneously acquire the expression of Nt5e (CD73), while Entpd1 (CD39) expression was not dramatically altered. Our studies implicate protective roles of CD73hi adipose Tregs and offer new therapeutic strategies against age-associated metabolic syndrome.

overall_design: mRNA profiles of LN Tregs (from 4-month old Foxp3Cre mice) and adipose Tregs (from 4-month-old Foxp3Cre, 4-month-old obese Foxp3Cre, 10-month old Foxp3Cre and 10-month-old Med23fl/fl;Foxp3Cre mice) were generated by deep sequencing, single experiment, using Illumina HiSeq2000.

first: Yangyang


title: The pioneer transcription factors Foxa1 and Foxa2 regulate alternative RNA splicing during positive selection in the thymus

status: Public on Jul 13 2021

first: Tessa

last: Crompton

address: 30 Guilford Street

department: Immunobiology

institute: UCL

summary: Here we show that the pioneer transcription factors Foxa1 and Foxa2 are required to regulate RNA splicing during positive selection at the transition from CD4+CD8+ double positive (DP) to single positive (SP thymocyte).  Conditional deletion of both Foxa1 and Foxa2 from DP thymocytes led to a partial arrest at the transition from DP to SP, with a reduction in the number of cells undergoing positive selection, and reduced the number and maturation of both CD4SP and CD8SP populations, with a reduction in peripheral naïve CD4+ T-cells.  Foxa1 and Foxa2 were required for physiological levels of expression of several genes which encoding splicing factors (Mbnl1, Sf3b1, Hnrnpa1) in cells undergoing positive selection.  Within the positively selecting CD69+DP cells, alternative RNA splicing was dysregulated leading to more than 859 significantly differentially expressed exons compared to control, with many genes important for T cell development and for RNA splicing showing differentially used exons.

overall_design: Thymocyte suspensions were prepared from adult (6 week old) Foxa1fl/flFoxa2fl/fl.CD4Cre-tg+ (Foxa1/2coKO) and Foxa1fl/flFoxa2fl/fl.Cre- (control) thymus.  Cells were stained with anti-CD69, anti-CD4 and anti-CD8 and facs sorted to purify CD69+CD4+CD8+ cells.  RNA was extracted from these CD69+CD4+CD8+ cells.

first: Ching-In

last: Lau


title: Golden hamster piRNAs are essential for early development and establishment of spermatogonia [hamster_oocyte.small_RNAseq]

city: Prague

first: Filip

last: Horvat

address: Videnska 1083

country: Czech Republic

institute: Institute of Molecular Genetics of the ASCR

summary: PIWI-associated RNAs (piRNAs) support germline by suppressing retrotransposons and genes. In mice, piRNAs are essential for spermatogenesis but not oogenesis. To test how this applies to other mammals, we deleted Mov10l1 helicase in golden hamsters whose piRNA pathway configuration is closer to other mammals than that of mice. Mov10l1–/– male hamsters showed impaired establishment of spermatogonia accompanied by transcriptome dysregulation and surge of MYSERV retrotransposon expression. The rare viable spermatogenic cells showed meiotic failure phenotype like Mov10l1–/– mice. Female Mov10l1–/–hamsters were sterile due to post-meiotic loss of developmental competence in zygotes. Unique phenotypes of Mov10l1–/– hamsters demonstrate adaptive nature of piRNA-mediated control of genes and retrotransposons, which enables confronting emerging genomic threats or acquiring new physiological roles.

overall_design:
Bulk small RNA-seq of 3 golden hamster GV oocyte samples:
- 2x WT
- 1x Mov10l1 KO

first: Petr

last: Svoboda

first: Radek

last: Malik

first: Zuzana

last: Loubalova


title: Murine Norovirus (MNV) infection results in anti-inflammatory response downstream of amino acids depletion in macrophages.

city: Guildford

first: Nicolas

last: Locker

state: Surrey

address: Stag Hill

department: FHMS

institute: University of Surrey

summary:
Purpose: Characterisation of MNV induced host genetic reprogramming and translational control exerted over the host response by comparison of available cytoplasmic transcripts and their association with polysomes.
Results: RNaseq data showed an inadequate response to MNV infection downstream of MDA5 activation coupled with a cellular response to a metabolic stress.  In particular, all 296 significantly regulated transcripts showed a strong correlation between a transcript availability in the cytoplasm and its recrutment to the polysomal fraction as addressed using the R package 'Anota2seq'.  Genome annotation analyses using Cytoscape highlighted the host response to the viral infection but showed a cluster of genes involved in response to stress and the related intrinsic apoptotic pathways suggesting a prevalence of a metabolic stress response over the innate immune response to the viral replication.
Conclusions: Our study provides the first understanding analysis of the host response to MNV at the level of available cytoplasmic mRNA and their recruitment to the actively translating fractions. It described an absence of translational control over the host innate immune response but highlight and inadequate NF-kB response correlating with an ectopic anti-inflammatory response to amino acid starvation.

overall_design: Methods: Cytoplasmic RNA were extracted  from cells infected with MNV or UV-irradiated MNV at different time post-infection (6 and 10hpi). For each sample,  half was purified directly (Total samples) and half loaded onto polysome gradient to obtain the RNA associated with the translation machinery (Polysomal samples) in triplicate.  Samples were enriched for polyA-tailed mRNA and sequenced on Illumina HiSeq4000, outputs quality checked via FastQC and mapped onto mouse genome (Gencode release M12 GRCm38.p5) using Hisat2 and to the Murine Norovirus genome MNV1-CW1 (DQ285629) coding sequences (ORF1: ABB90153.1, ORF2: ABB90154.1, ORF3: ABB90155.1) using  Bowtie. Genomic features of the host were defined using ‘Rsubread’ and in-built gene annotations for the mouse genome (NCBI RefSeq gene annotations Build 38.1). Host genomic features were then annotated using the R package ‘org.Mm.eg.db’, Ensembl and GenBank. The feature count matrix consisted of the set of Host and MNV feature count and genes annotated as rRNA, miRNA, snoRNA, snoRNA or scaRNA were filtered out of the feature count matrix.  Filtering of lowly expressed genes was performed after library size normalisation by keeping genes with at least 0.25 CPM in at least 25% of the samples in both Total and Polysomal fractions and differential expression addressed using the GLM implementation of EdgeR.  A final filtering criterion was applied to each individual significant gene using the genome visualisation tool Integrative Genome Viewer IGV hosted by the Broad Institute. qRT–PCR validation was performed using SYBR Green assays.


title: Golden hamster piRNAs are essential for early development and establishment of spermatogonia [hamster_testis.RNA_seq]

overall_design:
Bulk long RNA-seq of 21 golden hamster testis samples in five different ages:
- 0dpp: 2x WT, 2X Mov10l1 KO
- 9dpp: 3x WT, 2x Mov10l1 KO
- 13dpp: 2x WT, 2x Mov10l1 KO
- 21dpp: 2x WT, 2x Mov10l1 KO
- adult: 2x WT, 2x Mov10l1 KO


title: RNA-seq on NUT Carcinoma (NC) cell lines treated with DMSO, Panobinostat or IRBM6

first: Hitoshi

last: Shiota

summary: HDAC inhibitor induce transcriptional program driving cellular differentiation and growth arrest in NUT Carcinoma (NC) cell lines TC-797 and 10-15.

overall_design: RNA-seq on ribosomal RNA-depleded total RNA from NC cell lines TC-797 and 10-15 treated with DMSO, Panobinostat or IRBM6 for 4 hours.


title: Golden hamster piRNAs are essential for early development and establishment of spermatogonia.


title: Expression data from hippocampus in mouse brain

city: Tsukuba

first: Kazunori

last: Sasaki

address: 1-1-1 Tennodai

institute: AIST

summary:
Our microarray results showed there were up-regulated 28 genes and down-regulated 29 genes, which related depression and inflammatory response such as cytokine-cytokine receptor interaction, chemokine signaling pathway, dopaminergic synapse, glutamatergic synapse, GABAergic synapse, cholinergic synapse, and serotonergic synapse.
Among these genes, especially, higher hippocampal mRNA expression of transthyretin (Ttr), Zinc finger protein of the cerebellum 1 (Zic1), and Ectonucleotide pyrophosphatase/phosphodiesterase 2 (Enpp2) was found in kososan-administered defeated mice than water-administered defeated mice

overall_design: DNA microarray analysis was conducted using isolated RNAs extracted from the hippocampus as described previously (Sasaki et al., 2017). Double-stranded cDNA was synthesized from 100 ng of total RNA with the GeneAtlas 3' IVT Express Kit (Affymetrix Inc., Santa Clara, CA, USA). Biotin-labeled amplified RNA (aRNA) was synthesized using the GeneChip 3´ IVT Express Kit (Affymetrix Inc.). Purified aRNA was fragmented using the GeneAtlas 3´ IVT Express Kit and hybridized for 16 h at 45 °C using the GeneChip Mouse Genome 430 PM microarray (Affymetrix Inc.). The chip was washed and stained in the Gene Atlas Fluidics Station 400 (Affymetrix Inc.), and the resulting image was scanned using the GeneAtlas Imaging Station (Affymetrix Inc.). Data analysis was performed using the Affymetrix expression console (http://www.affymetrix.com) and the online data tool DAVID (Database for Annotation, Visualization and Integrated Discovery, v6.8, National Institute of Allergy and Infectious Diseases, NIH, USA, https://david.ncifcrf.gov/) (Huang et al., 2007a; Huang et al., 2007b). Among the clustered genes, the genes that contain the gene ontology concerning depression and inflammatory response (e.g., cytokine-cytokine receptor interaction, chemokine signaling pathway, dopaminergic synapse, glutamatergic synapse, GABAergic synapse, cholinergic synapse, and serotonergic synapse) were selected using DAVID tool and manually.


title: Golden hamster piRNAs are essential for early development and establishment of spermatogonia [hamster_testis.small_RNAseq]

overall_design:
Bulk small RNA-seq of 13 golden hamster testis samples in three different ages:
- 9dpp: 3x WT, 2x Mov10l1 KO
- 13dpp: 2x WT, 2x Mov10l1 KO
- 21dpp: 2x WT, 2x Mov10l1 KO


title: SLFN11 captures cancer-immunity interactions associated with platinum sensitivity in high-grade serous ovarian cancer

first: Joshua

last: Armenia

address: 1 Francis Crick Avenue

institute: AstraZeneca

summary:
Large independent analyses on cancer cell lines followed by functional studies have identified Schlafen 11 (SLFN11), a putative helicase, as the strongest predictor of sensitivity to DNA-damaging agents (DDA), including platinum. However, its role as a prognostic biomarker is undefined, partially due to the lack of validated methods to score SLFN11 in human tissues.
Here, we implemented a pipeline to quantify SLFN11 in human cancer samples. By analyzing a cohort of high-grade serous ovarian carcinoma (HGSOC) specimens  prior platinum-based chemotherapy treatment, we show, for the first time, that SLFN11 density in both the neoplastic and microenvironmental components was independently associated with favorable outcome.
We observed SLFN11 expression in both infiltrating innate and adaptive immune cells, and analyses in a second, independent, cohort revealed that SLFN11 is associated with immune activation in HGSOC. We found that platinum treatments activated immune-related pathways in ovarian cancer cells in a SLFN11-dependent manner, representative of tumor-immune transactivation.
Moreover, SLFN11 expression was induced in activated, isolated, immune cell subpopulations, hinting that SLFN11 in the immune compartment may be an indicator of immune transactivation.   In summary, we propose SLFN11 is a dual biomarker capturing simultaneously interconnected immunological and cancer-cell-intrinsic functional dispositions associated with sensitivity to DDA treatment.

overall_design: DU145 wild type and two different SLFN11 knockout clones were treated for 0, 6 or 24 hours with 100 nM gemcitabine

first: Claudia

last: Winkler

first: Matthew

last: King

first: Julie

last: Berthe

first: Domenico

last: Ferraioli

first: Anna

last: Garuti

first: Federica

last: Grillo

first: Jaime

last: Rodriguez-Canales

first: Lorenzo

last: Ferrando

last: Chopin

first: Isabelle

last: Ray-Coquard

first: Oona

last: Delpuech

first: Davide

last: Bedognetti

first: Alberto

last: Ballestrero

first: Elisabetta

last: Leo

first: Gabriele

last: Zoppoli


title: Investigation of the transcriptomic response of Atlantic salmon (Salmo salar) exposed to Neoparamoeba perurans.

city: Galway

first: Anita

middle: Mary

last: Talbot

state: Galway

address: Dublin Road Campus

department: Marine and Freshwater Research Centre

country: Ireland

institute: Galway-Mayo Institute of Technology

summary:
Purpose: The aim of this study was to RNA-sequencing (RNA-seq) to investigate further the gill transcriptome during the early stage of infection, prior to the appearance of mucoid lesions on the gills of  amoebic gill disease (ADG)-affected fish.
Methods: This study investigated the gill transcriptomic profile of pre-clinical AGD using RNA-sequencing (RNA-seq) technology. RNA-seq libraries generated at 0, 4, 7, 14 and 16 days post inoculation (dpi)  RNA-seq data was validated using real-time, quantitative PCR (qPCR) analysis.
Results: This study identified 29,357 Differentially Expressed Genes (DEGs) over the course of 16 days follwing exposure to N.perurans , the caustative agent of Amoebic gill disease.  With many genes differentially regualted at more tehn one time point, the number of individual gene identified as down-regulated was 8,524 and up-regulated was 10,826. DEGs mapped to 224 Gene Ontology (GO) terms, 27 reference pathways in the Kyoto Encyclopedia of Genes and Genomes (KEGG) and 15 Reactome Gene Sets.
Conclusions:Molecular diagnostics and histopathology, but not gill scores, provided an accurate record of disease progression during early stage onset of AGD, prior to evidence of clinical signs on the gill. At 7 days post infection (dpi), there was evidence of innate immune response activation and a concomitant immune suppression involving signalling pathways for cytokines, specifically interleukins.

overall_design: Atlantic salmon were exposed to N.perurans, the causative agent of AGD with samples collected over time five timepoints: 0, 4, 7, 14 and 16 dpi. There were 6 fish control and AGD-affected fish sampled at each  of teh four time points. Total RNA was extracted from gill tissue. 30 RNA-seq libraries were generated. The RNA-seq data was validated using qPCR. Differential gene expression was identified by comparing the individaul libraries at each point with the T0 control (T0_GCRL1 to T0_GCRL6).

first: Eugene

last: MacCarthy


title: Global analysis of the specificities and targets of endoribonucleases from E. coli toxin-antitoxin systems

first: Peter

last: Culviner

address: 677 Huntington Ave

department: IID

institute: Harvard School of Public Health

summary: The purpose of this study was to identify the RNA cleavage targets and specificities of 9 endoribonuclease toxins from toxin-antitoxin systems in E. coli MG1655.

overall_design: This study includes 80 paired-end RNA-Seq samples. The libraries were generated from E. coli cells expressing 9 different toxins or an empty vector from an arabinose inducible promoter. The study involved 4 sets of libraries with biological duplicates of each toxin or empty vector sample for each condition totaling 20 samples for each condition. For the first condition, toxins were induced for 5 minutes and libraries were rRNA depleted and not fragmented prior to reverse transcription. These libraries were intended to identify RNA 5’-ends generated by endoribonuclease toxin cleavage. In the second condition, toxins were induced for 5 minutes and libraries were rRNA depleted and fragmented for 3 minutes prior to reverse transcription. These libraries were intended to quantify cleavage at sites identified in the first condition by measuring the loss of signal in a toxin sample relative to the empty vector control. In the third condition, toxins were induced for 5 minutes and libraries were rRNA depleted, not fragmented, and subjected to a size selection to selectively sequence cDNA arising from tRNA. These libraries were intended to determine how toxin expression altered tRNA levels. In the fourth condition, toxins were induced for 30 minutes and libraries were not rRNA depleted and were not fragmented prior to reverse transcription. These libraries were intended to determine if rRNA is cleaved or rRNA maturation is altered by toxin expression.


title: BATF and IRF4 cooperate to counter exhaustion in tumour-infiltrating CAR T cells (RNA-seq)

city: La Jolla

first: Anjana

last: Rao

address: 9420 Athena Circle

department: Signaling and Gene Epression

institute: La Jolla institute

summary: Cooperative interactions among transcription factors are essential for gene transcription. We previously showed that NFAT and AP-1 (Fos-Jun) transcription factors cooperate to promote the effector functions of T cells, but that under conditions where it is unable to cooperate with AP-1, NFAT imposes a negative feedback programme of T cell hyporesponsiveness (“exhaustion”). Here we show that BATF and IRF4 cooperate to counter T cell exhaustion. Overexpression of Batf in CD8+ 42 T cells expressing a chimeric antigen receptor (CAR) promoted the survival and expansion of tumour-infiltrating CAR T cells, increased their production of effector cytokines, decreased their expression of inhibitory receptors and the exhaustion-associated transcription factor TOX, and led to the generation of long-lived memory T cells that controlled tumour recurrence. These responses were dependent on the BATF-IRF interaction, since cells expressing a Batf mutant unable to interact with Irf4 did not survive in tumours and did not effectively delay tumour growth. We suggest that BATF overexpression is a therapeutically viable option for improving the anti-tumour responses of CAR TILs, by skewing their phenotypes and transcriptional profiles away from exhaustion and towards increased effector function.

overall_design:
For transcriptional profiling using RNA-seq, 10,000 cells were sorted from the Live/Dead dye-negative CD8+Thy1.1+GFP+ population of the isolated tumor infiltrating lymphocytes or cultured CD8+ T cells. The cells were resuspended in FACS buffer and filtered with a 70 uM filter before sorting.
Gene expression with RNA-seq for CAR expressing CD8 T cells in CD8 T cells after in vitro culture after Re-Stimulation for 6h wwith CD3 & CD28

first: Hyungseok

last: Seo

first: Edahi

last: González-Avalos

first: Patrick

last: Hogan


title: Golden hamster piRNAs are essential for early development and establishment of spermatogonia [hamster_oocyte.bisulfite_seq]

first: Filip

last: Horvat

overall_design:
Bisulfite sequencing of 3 golden hamster GV oocyte samples:
- 1x WT
- 1x Mov10l1 HET
- 1x Mov10l1 KO

first: Petr

last: Svoboda

first: Radek

last: Malik

first: Zuzana

last: Loubalova


title: BATF and IRF4 cooperate to counter exhaustion in tumour-infiltrating CAR T cells (ATAC-seq)

overall_design: For transcriptional profiling using ATAC-seq, 50,000 cells were sorted from the Live/Dead dye-negative CD8+Thy1.1+GFP+ population of the isolated tumor infiltrating lymphocytes or cultured CD8+ T cells. The cells were resuspended in FACS buffer and filtered with a 70 M filter before sorting.


title: Single cell analysis of Dach1 overexpressing endothelial cells

city: Stanford

first: Kristy

last: Red-Horse

address: 318 Campus Drive

department: Biology

institute: Stanford Univeristy

summary: Overexpression of the transcriptional regulator Dach1 can induce coronary artery specification in embryonic mice. Here, we used single cell RNA sequencing to determine how Dach1 overexpression affects global gene expression to causes changes in arterial development. E15.5 control and endothelial specific Dach1 overexpression hearts were collection and FACS sorted to capture endothelial cells. 10x single cell libraries were then created and sequenced using Illumina high throughput sequencing. After identifying differentially expressed genes and assigning the cells to unique clusters, we found that the population of cells collected from Dach1 overexpression mice had a higher percentage of artery cells. Additionally, Dach1 overexpression upregulated the expression of artery genes and downregulated transcripts related to lipid metabolism in select clusters.

overall_design: scRNA seq from control and Dach1 overexpression mouse coronary endothelial cells

first: Brian

last: Raftrey

first: Ian

last: Williams


title: Excessive O-GlcNAcylation causes heart failure and sudden death

status: Public on Jul 10 2021

city: Baltimore

first: Mark

last: Anderson

state: MD

address: Rangos 471, 855 N. Wolfe Street

department: Department of Medicine

institute: Johns Hopkins School of Medicine

summary: Heart failure is a leading cause of death worldwide, and failing heart muscle is marked by increased O-GlcNAcylation (OGN).  It is unknown if excessive OGN contributes to cardiomyopathy and heart failure. OGN modifies serines and threonines, total OGN levels follow cellular nutrient and metabolic flux in addition to net activity of O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA).  We developed new transgenic mouse models with myocardial delimited over-expression of OGA and OGT, and found that OGT transgenic mice developed severe cardiomyopathy and premature death.  In contrast, OGA transgenic hearts had normal function, but were resistant to pathological stress. Interbreeding OGA transgenic mice rescued cardiomyopathy and premature death in OGT transgenic mice. RNA Seq and functional studies highlighted disrupted metabolism in hearts from OGT transgenic mice that was rescued by OGA transgenic interbreeding.  Here we show excessive OGN causes cardiomyopathy, identify gene programs responsive to pathological OGN, and suggest attenuation of OGN may be an effective therapy for heart failure.

overall_design: This poly(A+) transcriptome profiling study examined the cardiac transcriptional profiles of adult wildtype, transgenic OGA and OGT mice, and OGA x OGT double transgenic mice.

first: Priya

last: Umapathi

first: Partha

middle: S

last: Banerjee

first: Natasha

last: Zachara

first: Neha

last: Abrol

first: Qinchuan

last: Wang

first: Olurotimi

middle: O

last: Mesubi

first: Elizabeth

middle: D

last: Luczak

first: Yuejin

last: Wu

first: Jonathan

last: Granger

first: Liliana

last: Florea

first: Conover

first: Gerald

last: Hart


title: Golden hamster piRNAs are essential for early development and establishment of spermatogonia [hamster_oocyte.RNA_seq]

first: Filip

last: Horvat

overall_design:
Bulk long RNA-seq of 7 golden hamster GV oocyte samples:
- 2x WT
- 2x Mov10l1 HET
- 3x Mov10l1 KO

first: Petr

last: Svoboda

first: Radek

last: Malik

first: Zuzana


title: MeRIP sequencing analysis of BJAB cell line infected with EBV

city: Changsha

first: Jian

last: Ma

state: Hunan

address: 110  Xiangya  Road

department: Cancer  Research Institute

institute: Central South University

summary:
Purpose: Evaluation of the m6A modification of EBV and BJAB transcripts during EBV infection
Methods: Human B lymphoma cell line BJAB was uninfected or infected with EBV for 24 hours. Total RNA from each sample were extracted. Intact mRNA was isolated from total RNA samples and then chemically fragmented to 100-nucleoside-long fragments. m6A methylated mRNAs were immunoprecipitated with anti-N6-methyadenosine (m6A) antibody (a part of the fragmented mRNAs was kept as input). Both m6A enriched mRNAs and input mRNAs were concentrated for RNA-seq library construction. Sequencing was performed using an Illumina HiSeq 4000.
Results: EBV EBNA2 and BHRF1 transcripts were m6A modified and m6A modification of BJAB transcripts changed during EBV infection.
Conclusions: Our study found that some EBV transcripts were m6A modified during EBV infection and EBV infection changed m6A modification profiles of BJAB transcripts.

overall_design: Three independent biological replicates were plated and uninfected or infected with EBV in parallel.

first: Xiang

last: Zheng

first: Jia


title: An epigenetic program established by gene – environment interactions initiates pancreatic carcinogenesis [ATACSeq_shBrd4]

first: Yu-Jui

last: Ho

state: New York

address: 417 E 68th St

department: Cancer Biology & Genetics Program

institute: Memorial Sloan Kettering Cancer Center

summary: Damaged tissues have increased risk of cancer development through poorly understood mechanisms. In the pancreas, tissue damage collaborates with activating mutations in the Kras oncogene to dramatically accelerate the formation of early neoplastic lesions and ultimately pancreatic cancer. By integrating genomics, single-cell chromatin assays and spatiotemporally-controlled functional perturbations in autochthonous mouse models, we show that the combination of Kras mutation and tissue damage promotes a unique chromatin state in the pancreatic epithelium that distinguishes neoplastic transformation from normal regeneration and is selected for throughout malignant evolution. This cancer-associated epigenetic state emerges within 48 hours of pancreatic injury, and involves large-scale chromatin accessibility changes that contribute to the early dysregulation of genes defining human pancreatic cancer. Among the genes most rapidly activated by tissue damage in the pre-malignant pancreatic epithelium is the alarmin cytokine IL33, which cooperates with mutant Kras in unleashing the epigenetic remodeling program of early neoplasia and neoplastic transformation in the absence of injury. Collectively, our study demonstrates how gene-environment interactions can rapidly produce gene regulatory programs that dictate early neoplastic commitment and provides a molecular framework for understanding the interplay between genetics and environmental cues in cancer initiation.

overall_design: Characterization of the chromatin accessibility (ATAC-seq) profiles of bulk lineage-traced Kras-mutant pancreatic epithelial cells expressing shRen.713 or shBrd4.552, isolated by FACS-sorting from pancreata of on dox KCsh mice subjected to caerulein treatment (Kras*+Injury condition).

first: Direna

last: Curbelo

first: Richard

middle: P

last: Koche

first: Scott

last: Lowe


title: Modeling Alcohol-induced Neurotoxicity using Human Induced Pluripotent Stem Cell-derived Three-Dimensional Cerebral Organoids

city: Milwaukee

first: Thiago

last: Arzua

state: WI

address: 8701 W Watertown Plank Rd

institute: Medical College f Wisconsin

type:
Non-coding RNA profiling by array
Expression profiling by array

summary:
We performed transcriptomic studies using microarray assays and conducted bioinformatics analyses to identify signaling pathways associated with ethanol-induced brain injury in cerebral organoids
A human message RNA (mRNA) Expression Microarray assay (V4.0) and data analysis services were provided by Arraystar Inc. (Rockville, MD) to assess and compare the global abundance of 17,195 mRNAs across iPSC-derived 2-month cerebral organoids treated with and without ethanol.

overall_design:
After a 6 hours exposure to either 50 mM of ethanol or a control (normal media), n=4, RNA was isolated from cerebral organoids converted to cDNA, synthesized from RNA via reverse transcription, and hybridized to Arraystar’s custom mRNA probes.
contributor Arraystar Inc.

first: Yasheng

last: Yan

first: Congshan

last: Jiang

first: Sarah

last: Logan

first: Reilly

last: L. Allison

first: Clive

last: Wells

first: Suresh

last: Kumar

last: Schäfer

first: Xiaowen

last: Bai


title: Next Generation Sequencing Facilitates Quantitative Analysis of the MDM2i and WIPi Combination Therapy in Intrahepatic Cholangiocarcinoma Transcriptomes

status: Public on Jul 09 2021

city: Taoyuan

first: Chiao-Ping

last: Chen

address: No.259, Wenhua 1st Rd., Guishan Dist.

institute: Chang Gung University

summary:
Purpose: Next-generation sequencing (NGS) has revolutionized the systems-based analysis of cellular pathways. The purpose of this study is to compare the expression profiles of single and combination treatment of MDM2i and WIP1i in RBE cells by NGS-derived transcriptome profiling (RNA-seq).
Methods: RBE mRNA profiles of HDM201 (MDM2i) and/or GSK2830371 (WIP1i) treatment for 6 or 24 hours were generated by deep sequencing, in triplicate, using Illumina NextSeq sequencer. The sequence of each read was trimmed based on the quality score (Q30), and read lengths less than 45 bp were dis-carded prior to further analysis. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods:  STAR aligner followed by ANOVA.
Results: We mapped about 30 million sequence reads per sample to the RBE transcriptomes after HDM201 and/or GSK2830371 treatment. The differentially expressed genes were selected for more than 2-fold changes and p-values for differences in expression of less than 0.05. From four different comparisons, HDM201 alone and in combination with GSK2830371 for 6 h and 24 h, 26 consistently altered genes were identified (21 upregulated genes and 5 downregulated genes).
Conclusions: Our study represents the detailed analysis of MDM2i and/or WIPi therapy in intrahepatic cholangiocarcinoma transcriptomes. Differentially expressed genes from combination treatment were enriched for genes involved in p53-dependent signal transduction pathways, involved in the DNA damage response, response to stress, and regulation of apoptotic processes. In addition, differentially expressed genes, analyzed using the KEGG pathway da-tabase, were enriched for genes involved in platinum drug resistance and the p53 signaling pathway and gene ontology database for protein-protein interaction using an adjusted p-values less than 0.01.

overall_design: The mRNA profiles of MDM2i and/or WIP1i treatment for 6 or 24 hours in RBE

first: Chiao-En

last: Wu


title: BATF and IRF4 cooperate to counter exhaustion in tumour-infiltrating CAR T cells

first: Anjana

last: Rao

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: The synergistic anti-tumor activity of EZH2 inhibitor SHR2554 and HDAC inhibitor chidamide through ORC1 reduction of DNA replication process in diffuse large B cell lymphoma

status: Public on Jul 12 2021

city: Beijing

first: Jun

last: Zhu

address: No.52 Fucheng Road, Haidian District

institute: Department of Lymphoma, Peking University Cancer Hospital & Institute

summary: Background: Upregulation of H3K27me3 induced by EZH2 overexpression or somatic heterozygous mutations were implicated in lymphomagenesis. It has been demonstrated that several EZH2-target agents have notable therapeutic effects in EZH2-mutant B-cell lymphoma patients. Here we present a novel highly selective EZH2 inhibitor SHR2554 and possible combination strategy in diffuse large B cell lymphoma (DLBCL); Methods: Cell proliferation, cycle and apoptosis were analyzed by Cell Titer-Glo Luminescent Cell Viability Assay and flow cytometry. Western Blot was used to detect the regulatory protease in related signaling pathways and RNA-seq was conducted to assess transcriptome changes. Finally, CDX and PDX models were used to evaluate the synergistic anti-tumor effects of the combination in vivo; Results: The novel EZH2 inhibitor SHR2554 could inhibited proliferation, induced G1 phase arrest in EZH2-mutant DLBCL cell lines. The combination of EZH2 inhibitor SHR2554 with histone deacetylase (HDAC) inhibitor chidamide (hereafter referred as HBI8000) exerted synergistic anti-proliferative activity in vitro and in vivo. Gene expression profile analysis revealed dramatic inhibition of DNA replication process in combined treatment; Conclusions: SHR2554, a potent highly selective small molecule inhibitor of EZH2, inhibited EZH2-mutant DLBCL more significantly in vitro and in vivo. The combination of HDAC inhibitor HBI8000 with EZH2 inhibitor SHR2554 exhibited dramatically anti-tumor activity in both mutant and wild-type DLBCL, which may become potential therapeutic modality for the treatment of DLBCL patients.

overall_design: RNA sequencing in 2  DLBCL cell lines with SHR2554  and/or HBI8000 treatment .

first: Xing

last: Wang

first: Dedao

first: Ning

last: Ding

first: Yuqin

last: Song


title: Macrophage-regulatory T cell interactions promote type 2 immune homeostasis through Resistin-like molecule α

city: Riverside

first: Meera

middle: G

last: Nair

address: 3401 Watkins Drive

department: Division of Biomedical Sciences

institute: University of California Riverside

summary: RELMα is a small, secreted protein expressed by T helper type 2 (Th2) cytokine activated “M2” macrophages in helminth infection and allergy. At steady state and in response to Th2 cytokines, RELMα is highly expressed by peritoneal macrophages. We generated RELMα TdTomato (Td) reporter/knockout (RΕLMαTd/Td) mice. RELMαTd/Td mice had increased IL-4 induced peritoneal macrophage responses and splenomegaly compared to RELMα+/+ mice. Nanostring data indicated that RELMαTd/Td peritoneal macrophages were more proliferative and activated than RELMα+/+ macrophages, with increased genes associated with T cell response, growth factor and cytokine signaling, but decreased genes associated with differentiation and maintenance of myeloid cells.This studies identify a new RELMα regulatory pathway whereby RELMα-expressing macrophages directly sustain Treg proliferation to limit Th2 inflammatory responses.

overall_design: 11 samples are included, 2 from PBS injected Rα+/+, 2 from PBS injected RαΤd/Td, 3 from IL4c injected Rα+/+ and 4 from IL4c injected RαTd/Td. Mice injected i.p. with mouse IL-4c or PBS (vehicle control) on days 0 and 2 and peritoneal cells were recovered at day 4. Peritoneal macrophages (CD11b+F4/80+) were sorted with the MoFlo Astrios cell sorter then anaylzed with Nanostring.

first: Jiang

last: Li

first: Sang

last: Kim


title: BATF and IRF4 cooperate to counter exhaustion in tumour-infiltrating CAR T cells (ChIP-seq)

overall_design:
ChiP-seq: Determination of BATF or IRF4 binding sites in CD8 T cells in vitro and in Re-Stimulated CD8 T cells in vitro
Chromatin Immunoprecipitation followed by Sequencing for CD8 T cells after in vitro culture and Re-Stimulation 6h

first: Hyungseok

last: Seo

first: Edahi

last: González-Avalos

first: Patrick

last: Hogan


title: An epigenetic program established by gene – environment interactions initiates pancreatic carcinogenesis [ATACSeq_rIL33]

first: Yu-Jui

last: Ho

overall_design: Characterization of the chromatin accessibility profiles (ATAC-seq) of bulk lineage-traced Kras-mutant pancreatic epithelial cells isolated by FACS-sorting from pancreata of KC (p48-Cre;RIK;LSLKrasG12D) mice treated with murine recombinant IL-33 (rIL-33) or PBS.

first: Direna

last: Curbelo

first: Richard

last: Koche

first: Scott

middle: W

last: Lowe


title: Modeling High-grade serous ovarian carcinoma using a combination of in vivo fallopian tube electroporation and CRISPR/Cas9-mediated genome editing

city: montreal

first: Yojiro

last: Yamanaka

state: Quebec

address: 1160 Pine Avenue West, room 416

department: Human Genetics

institute: McGill University (Goodman Cancer Research Centre)

summary: In this study we generated a mouse model of high-grade serous ovarian carcinoma by targeting tumor suppressor genes Brca1, P53 and Pten using CRISPR/Cas9. The addition of Lkb1 to this combination reduced tumor latency and changed the origin of the tumor from the fallopain tube to the ovarian surface epithelium. Tumors originating the ovarian surface had distinct copy number alterations, highlighting the impact of the cell-of-origin on susceptibility and tumor development. In addition using lineage tracing and single cell RNA sequnecing techniques we investigated ovarian cancer ascities and early metastasis in the peritoneal cavity.

overall_design: Whole genome sequencing of ovarian tumors and scRNA sequencing of ascties fluild isolated from a single mouse.

first: Katie

last: Teng

first: Matthew

middle: J

last: Ford


title: Next Generation Sequencing Facilitates Quantitative Analysis of Mock/MASCp36 infected mouse lung Transcriptomes

first: Rui-Ting

state: Beijing

address: No.20 Dongda Street, Fengtai District

department: State Key Laboratory of Pathogen and Biosecurity

institute: Beijing Institute of Microbiology and Epidemiology

summary: The ongoing SARS-CoV-2 pandemic has brought an urgent need for animal models to study the pathogenicity of the virus. Herein, we generated and characterized a novel mouse-adapted SARS-CoV-2 strain named MASCp36 that causes acute respiratory symptoms and mortality in standard laboratory mice. Particularly, this model exhibits age and gender related skewed distribution of mortality akin to severe COVID-19, and the 50% lethal dose (LD50) of MASCp36 was ~100 PFU in aged, male BALB/c mice. To characterize the transcriptional response to infection of MASCp36 in BALB/c mice (different age and gender), RNA-Seq analysis were performed using lung homogenates collected at 1 and 4 dpi.

first: Cheng-Feng

last: Qin

first: Hong-Jing

last: Gu


title: Aspects of the Neurospora crassa sulfur starvation response are revealed by transcriptional profiling and DNA affinity purification sequencing

city: Ithaca

first: Lori

last: Huberman

address: 236 Tower Rd.

department: Plant Pathology and Plant-Microbe Biology, School of Integrative Plant Science

institute: Cornell University

summary: Accurate nutrient sensing is important for rapid fungal growth and exploitation of available resources. Sulfur is an important nutrient source found in a number of biological macromolecules, including proteins and lipids. The model filamentous fungus Neurospora crassa is capable of utilizing sulfur found in a variety of sources from amino acids to sulfate. During sulfur starvation, the transcription factor CYS-3 is responsible for upregulation of genes involved in sulfur uptake and assimilation. Using a combination of RNA sequencing and DNA affinity purification sequencing, we performed a global survey of the N. crassa sulfur starvation response and the role of CYS-3 in regulating sulfur responsive genes. Along with genes known to be involved in sulfur metabolism, the CYS-3 transcription factor also directly activated the expression of a number of uncharacterized transporter genes, suggesting that regulating sulfur import is an important aspect of regulation by CYS-3. Additionally, CYS-3 directly regulated the expression of genes involved in mitochondrial electron transfer. During sulfur starvation, genes involved in nitrogen metabolism, such as amino acid and nucleic acid metabolic pathways, along with genes encoding proteases and nucleases that are necessary for scavenging nitrogen, were activated. Sulfur starvation also caused changes in the expression of genes involved in carbohydrate metabolism, such as those encoding glycosyl hydrolases. Thus, our data suggest a connection between sulfur metabolism and other aspects of cellular metabolism.

overall_design: mRNA profiles of wild type N. crassa cells and N. crassa cells with a cys-3 partial deletion exposed to media with 24uM sulfate and sulfur starvation for 4 hours. Each of the two strains (mutant and wild type control) was exposed to both sulfur conditions for 4 total experiments, each done in biological triplicate. There are a total of 12 samples.

first: Vincent

last: Wu

first: Ronan

middle: C

last: O'Malley

first: N

last: Glass


title: Fli1+ cells transcriptional analysis reveals LMO2-PRDM16 axis in angiogenesis

city: Edinburgh

first: Gianfranco

last: Matrone

address: 47 Little France Cresc

institute: University of Edinburgh

summary: Different GFP+ cell population were isolated from Tg(fli1:EGFP) zebrafish embryos. Novel roles of epigenetic modifiers in endothelial differentiation were identified.

overall_design: Each sample has been run in duplicate, with 2 files per each run

first: Bo

last: Xia

first: Kaifu

last: Chen

first: Martin

middle: A

last: Denvir

first: Andrew

last: Baker

first: John

last: Cooke